In this proposal, we will use molecular genetic interventions in conjunction with other treatment modalities to maximally inhibit viral replication and optimize immune function in pediatric and adult AIDS. The goal of this program will be to perform preclinical and clinical studies in cell culture, animal models, and patients to develop optimal combinations of gene therapy, antiviral drugs and immunostimulation for HIV infection. The program develops on cross-disciplinary efforts of the five independent investigators in this area, which has provided the basis for previous productive collaborations among the participating basic scientists and clinicians. In Project 1, Dr. Nabel will optimize the composition of gene transfer vectors and include antiviral genes targeted toward different phases of the virus life cycle, targeting chemokine receptors or other afferent steps, in addition to Rev M10, directed toward the efferent phase. Vector production systems with several advantages for human studies will be optimized and clinical protocols which examine the effect of combinations of gene therapy, drug treatment and immunostimulation will be performed. Dr. Kohn (Project 2) will implement a human gene therapy protocol to perform gene transfer into hematopoietic cells and evaluate its efficacy in pediatric AIDS, where such an approach appears more clinically indicated. A lentiviral based vector system which will target T cells and early hematopoietic cells will be developed in Project 3 (Dr. Nolan), and will be analyzed in preclinical models and potential in clinical studies, in collaboration with Projects 1 and Projects 2. In Project 4, Dr. Littman will produce knockouts or mouse strains with chemokine mutants known to bind and inhibit CCR5 and CXCR4 function. Their effects in transgenic models of chemokine receptor antagonism and knockout will be studied, complementing efforts to develop gene therapy vectors targeted towards these gene products (Project 1). Two core facilities will provide support for vector production, virologic studies and immune analyses: a BSL3 lab/GMP facility will assess the protective effect of these regulated genes and prepare vectors for clinical studies, and an Immunology Core will analyze immune responses to vectors and recombinant genes in transduced cells and assist in the analysis of immune function in chemokine receptor defective mice (Project 4). Pilot transduced cells and assist in the analysis of immune function in chemokine receptor defective mice (Project 4). Pilot studies will be performed which can be expanded in the future to Phase II/III clinical studies. This proposal represents a multi-disciplinary approach for the ration development of novel, combination treatments for HIV infection to further our understanding of the pathophysiology and treatment responses to AIDS.